Non-viral gene delivery agent comprising lipopeptide (LP) compounds
Non-viral nucleic acid delivery agents and methods for the delivery or transfer of nucleic acid molecules to target cells are disclosed. The agents comprise a complex of a nucleic acid cargo for delivery, one or more lipopeptide compound, and one or more polymeric charge-neutralising agent, and the complex is in the form of a particle with substantially neutral or negative surface charge. The agents and methods may be useful in a variety of applications such as therapies (including gene therapies and nucleic acid vaccinations) for diseases and medical disorders.
1. A non-viral nucleic acid delivery agent comprising a complex of:
(i) a nucleic acid cargo for delivery to a cell;
(ii) one or more cationic lipopeptide compound; and
(iii) one or more polymeric charge-neutralising agent selected from the group consisting of poly-amino acid polymers, copolymers of polyethylene glycol (PEG) and a poly-amino acid polymer, and copolymers of poly(acrylic acid);
wherein said complex is in the form of a particle with substantially neutral or negative surface charge, and the formation of said complex is conducted in the aqueous phase.
2. The delivery agent of claim 1 , wherein the nucleic acid cargo is a DNA molecule which encodes a protein(s), oligopeptide(s) or peptide(s).
3. The delivery agent of claim 1 , wherein the nucleic acid cargo is an mRNA molecule which encodes a protein(s), oligopeptide(s) or peptide(s).
4. The delivery agent of claim 1 , wherein the nucleic acid cargo is a modRNA.
5. The delivery agent of claim 1 , wherein the lipopeptide compound is of the general formula I:
(I) R-L-peptide
wherein R is a linear or branched alkyl;
L is a linker group; and
the peptide is of any amino acid sequence comprising 2-15 amino acids, but with the proviso that at least two of the amino acids are independently selected from those with strongly basic/positively charged properties.
6. The delivery agent of claim 1 , wherein the lipopeptide compound is of the general formula II:
(II) R—CO-peptide
wherein
R is CH 3 —(CH 2 ) n —, where n is an integer in the range of 11 to 21, and the peptide is of any amino acid sequence comprising 2-15 amino acids, but with the proviso that at least two of the amino acids are independently selected from those with strongly basic/positively charged properties.
7. The delivery agent of claim 5 , wherein the peptide comprises 5-10 amino acids.
8. The delivery agent of claim 5 , wherein at least two of the amino acids of the amino acid sequence are independently selected from lysine (Lys), arginine (Arg) and histidine (His).
9. The delivery agent of claim 6 , wherein the lipopeptide compound is of the general formula: stearoyl-Cys-X 1 -X 2 -Lys-Lys-Lys (SEQ ID NO: 2), where X 1 and X 2 are any amino acids and may be the same or different.
10. The delivery agent of claim 6 , wherein the lipopeptide compound is of the general formula: stearoyl-X 0 -X 1 -X 2 -Lys-Lys-Lys, where X 0 is absent (X 1 -X 2 -Lys-Lys-Lys; SEQ ID NO:14) or any amino acid other than Cys (X 0 -X 1 -X 2 -Lys-Lys-Lys; SEQ ID NO:3), and X 1 and X 2 are any amino acids other than Cys and may be the same or different.
11. The delivery agent of claim 10 , wherein X 0 is Ser or Thr.
12. The delivery agent of claim 9 , wherein X 1 and X 2 are independently selected from Ala, Arg and His.
13. The delivery agent of claim 10 , wherein X 1 and X 2 are independently selected from Ala, Arg and His.
14. The delivery agent of claim 9 , wherein X 1 and X 2 are His.
15. The delivery agent of claim 10 , wherein X 1 and X 2 are His.
16. The delivery agent of claim 1 , wherein the polymeric charge-neutralising agent(s) is selected from poly(L-glutamate)-PEG copolymers (PLGA-PEG) or copolymers of poly(acrylic acid).
17. The delivery agent of claim 16 , wherein the polymeric charge-neutralising agent(s) is selected from block copolymers of poly(acrylic acid) and poly(hydroxypropyl methacrylamide) (polyHPMA), and block copolymers of poly(acrylic acid) and poly(2-hydroxyethyl methacrylamide) (polyHEMA).
18. The delivery agent of claim 1 , wherein the particle shows a zeta potential (ZP) surface charge in the range of −2 to 2 mV or −40 to −5 mV.
19. A method of delivering a nucleic acid molecule to a cell of a subject, said method comprising the steps of:
providing a non-viral nucleic acid delivery agent according to claim 1 ; and
delivering the non-viral nucleic acid delivery agent to said cell of the subject.
20. The method of claim 19 , wherein the non-viral nucleic acid delivery agent is delivered parentally to the subject.
21. A pharmaceutical composition comprising a non-viral delivery nucleic acid agent according to claim 1 in combination with a pharmaceutically acceptable carrier or excipient.
22. A method of producing a non-viral delivery nucleic acid agent according to claim 1 , said method comprising:
combining a nucleic acid cargo, one or more lipopeptide compound, and one or more polymeric charge-neutralising agent in the aqueous phase under conditions suitable for the formation of complexed particles.